scholarly journals Contractions transport exfoliated epithelial cells through the neonatal epididymis

Reproduction ◽  
2020 ◽  
Vol 160 (1) ◽  
pp. 109-116
Author(s):  
Daniela Weiser ◽  
Andrea Mietens ◽  
Beatrix Stadler ◽  
Davor Ježek ◽  
Gerhard Schuler ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Time Lapse ◽  
Neutral Endopeptidase ◽  
Neonatal Rat ◽  
Nuclear Antigen ◽  
Rat Epididymis ◽  
Luminal Cells ◽  
Pendular Movement ◽  
Spontaneous Contractions ◽  
Epididymal Duct

Contractions of the adult epididymal duct are well known in the context of sperm transport. Some reports also describe contractions of the epididymal duct during development, but data about their character, regulation and function are sparse. In the foetal human epididymis we found luminal cells and could identify them as exfoliated epithelial cells originating from the epididymis and not from testis by using antibodies against neutral endopeptidase as an epithelial epididymal duct marker. Exfoliated cells were also found in the epididymal duct after birth. Time-lapse imaging revealed directional transport of luminal cells in the neonatal rat epididymis interrupted by pendular movement. Spontaneous contractions were discovered in the neonatal epididymis and an association between these contractions and the transport of the luminal cells could be observed. Both, transport and spontaneous contractions, were affected significantly by substances known to contract (noradrenaline) or relax (the phosphodiesterase 5 inhibitor sildenafil) smooth muscle cells. Immunohistochemistry showed staining for the proliferation marker proliferating-cell-nuclear-antigen (PCNA) in cells of the ductal lumen of the neonatal rat epididymis indicating the extrusion of cells also during proliferation. Our data showed spontaneous contractions of the immature epididymal duct associated with the transport of exfoliated luminal cells before the first occurrence of sperm cells. Results suggest an important role including both (i) a mechanical place holder function of exfoliated luminal cells (ii) together with a novel idea of organized waste disposal of these cells during development.

Contact behaviour during the reassociation of dissociated epithelial cells in primary culture

1987 ◽  
Vol 88 (4) ◽  
pp. 521-526
Author(s):  
R.M. Brown ◽  
C.A. Middleton
Keyword(s):  
Epithelial Cells ◽  
Chick Embryo ◽  
Primary Culture ◽  
Cell Cultures ◽  
Corneal Epithelium ◽  
Time Lapse ◽  
Cell Types ◽  
Epidermal Cells ◽  
Isolated Cells ◽  
Contact Behaviour

The behaviour in culture of dissociated epithelial cells from chick embryo pigmented retina epithelium (PRE), corneal epithelium (CE) and epidermis has been studied using time-lapse cinematography. The analysis concentrated on the contact behaviour of 60 previously isolated cells of each type during a 24 h period starting 3.5 h after the cells were plated out. During the period analysed the number of isolated cells in cultures of all three types gradually decreased as they became incorporated into islands and sheets of cells. However, there were significant differences in behaviour between the cell types during the establishment of these sheets and islands. In PRE cell cultures, islands of cells developed because, throughout the period of analysis, collisions involving previously isolated cells almost invariably resulted in the development of a stable contact. Once having established contact with another cell these cells rarely broke away again to become reisolated. In contrast the contacts formed between colliding CE and epidermal cells were, at least initially, much less stable and cells of both these types were frequently seen to break away and become reisolated after colliding with other cells. Sheets and islands of cells eventually developed in these cultures because the frequency with which isolated cells become reisolated decreased with increasing time in culture. The possible reasons underlying the different behaviour of PRE cells, when compared with that of CE and epidermal cells, are discussed. It is suggested that the decreasing tendency of isolated CE and epidermal cells to become reisolated may be related to the formation of desmosomes.

Differential expression of neutral endopeptidase-24.11 (neprilysin) and endothelin-converting enzyme in human prostate cancer cell lines

2002 ◽  
Vol 103 (s2002) ◽  
pp. 314S-317S ◽  
Author(s):  
Badar A. USMANI ◽  
Ben HARDEN ◽  
Norman J. MAITLAND ◽  
Anthony J. TURNER
Keyword(s):  
Prostate Cancer ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Prostate Cancer Cell ◽  
Neutral Endopeptidase ◽  
Prostatic Hyperplasia ◽  
Converting Enzyme ◽  
Prostate Cancer Cell Lines ◽  
Endothelin Converting Enzyme ◽  
Androgen Independent

Neutral endopeptidase-24.11 (neprilysin; NEP/CD10) is a cell surface metallopeptidase expressed by prostatic epithelial cells that degrades various bioactive peptides including endothelin. Endothelin-converting enzyme (ECE), the key enzyme of endothelin biosynthesis, catalyses the final processing step in the pathway. Neuropeptide substrates of NEP, including endothelin, have been implicated in the growth of androgen-independent prostate cancer. We have surveyed the expression of NEP and ECE in a range of prostate cancer cell lines. Western analysis reveals that ECE-1 is expressed abundantly in all the malignant cell lines tested, except for LNCaP. In contrast, LNCaP cells express high levels of NEP, while NEP was not detected in PC-3, DU145 and other metastatic cell lines that were tested. Of the normal immortalized prostate epithelial cell lines, PNT1a shows equivalent amounts of NEP and ECE. PNT2-C2 shows poor NEP expression but an abundance of ECE. P4E6, by comparison, has low levels of both ECE and NEP. These differences in expression may render these cell lines useful in experimental models for future study. Benign prostatic hyperplasia primary epithelial cells express much higher levels of NEP than malignant primary epithelial cells, but neither show ECE expression. On the other hand, surrounding stromal cell populations have detectable ECE levels. An absence of ECE in malignant and benign prostatic hyperplasia cells of primary epithelial origin suggests an important role for stromal interaction and paracrine production of ECE within the host. The upregulation of ECE expression in metastatic cells in culture may be indicative of its role in metastatic progression. A differential profile of ECE and NEP could contribute to an abundance of mitogenic peptides aiding the progression of androgen-independent prostate cancer.

scholarly journals Castration causes an increase in lysosomal size and upregulation of cathepsin D expression in principal cells along with increased secretion of procathepsin D and prosaposin oligomers in adult rat epididymis

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0250454
Author(s):  
Lorena Carvelli ◽  
Andrea Carolina Aguilera ◽  
Leila Zyla ◽  
Laura Lucía Pereyra ◽  
Carlos R. Morales ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Complex Formation ◽  
Cathepsin D ◽  
Initial Segment ◽  
Lysosomal Enzymes ◽  
Sperm Maturation ◽  
Principal Cells ◽  
Adult Rat ◽  
Rat Epididymis ◽  
Lysosomal Proteins

In the epididymis, lysosomal proteins of the epithelial cells are normally targeted from the Golgi apparatus to lysosomes for degradation, although their secretion into the epididymal lumen has been documented and associated with sperm maturation. In this study, cathepsin D (CatD) and prosaposin (PSAP) were examined in adult epididymis of control, and 2-day castrated rats without (Ct) and with testosterone replacement (Ct+T) to evaluate their expression and regulation within epididymal epithelial cells. By light microscope-immunocytochemistry, a quantitative increase in size of lysosomes in principal cells of Ct animals was noted from the distal initial segment to the proximal cauda. Androgen replacement did not restore the size of lysosomes to control levels. Western blot analysis revealed a significant increase in CatD expression in the epididymis of Ct animals, which suggested an upregulation of its expression in principal cells; androgens restored levels of CatD to that of controls. In contrast, PSAP expression in Ct animals was not altered from controls. Additionally, an increase in procathepsin D levels was noted from samples of the epididymal fluid of Ct compared to control animals, accompanied by an increased complex formation with PSAP. Moreover, an increased oligomerization of prosaposin was observed in the epididymal lumen of Ct rats, with changes reverted to controls in Ct+T animals. Taken together these data suggest castration causes an increased uptake of substrates that are acted upon by CatD in lysosomes of principal cells and in the lumen by procathepsin D. These substrates may be derived from apoptotic cells noted in the lumen of proximal regions and possibly by degenerating sperm in distal regions of the epididymis of Ct animals. Exploring the mechanisms by which lysosomal enzymes are synthesized and secreted by the epididymis may help resolve some of the issues originating from epididymal dysfunctions with relevance to sperm maturation.

scholarly journals Functional Calsequestrin-1 Is Expressed in the Heart and Its Deficiency Is Causally Related to Malignant Hyperthermia-Like Arrhythmia

Circulation ◽  
2021 ◽  
Author(s):  
Zhipeng Sun ◽  
Luqi Wang ◽  
Lu Han ◽  
Yue Wang ◽  
Yuan Zhou ◽  
...  
Keyword(s):  
Heart Rate ◽  
Ventricular Tachycardia ◽  
Malignant Hyperthermia ◽  
Ex Vivo ◽  
Heat Stroke ◽  
Time Lapse ◽  
Underlying Mechanism ◽  
Neonatal Rat ◽  
Polymorphic Ventricular Tachycardia

Background: Calsequestrins (Casqs), comprising the Casq1 and Casq2 isoforms, buffer Ca 2+ and regulate its release in the sarcoplasmic reticulum (SR) of skeletal and cardiac muscle, respectively. Human inherited diseases associated with mutations in CASQ1 or CASQ2 include malignant hyperthermia/environmental heat stroke (MH/EHS) and catecholaminergic polymorphic ventricular tachycardia. However, patients with an MH/EHS event often suffer from arrhythmia for which the underlying mechanism remains unknown. Methods: Working hearts from conventional ( Casq1 -KO) and cardiac-specific ( Casq1 -CKO) Casq1 knockout mice were monitored in vivo and ex vivo by electrocardiogram and electrical mapping, respectively. MH was induced by 2% isoflurane and treated intraperitoneally with dantrolene. Time-lapse imaging was used to monitor intracellular Ca 2+ activity in isolated mouse cardiomyocytes or neonatal rat ventricular myocytes (NRVMs) with knockdown, over-expression or truncation of the Casq1 gene. Conformational change in both Casqs was determined by crosslinking Western blot analysis. Results: Like MH/EHS patients, Casq1 -KO and Casq1 -CKO mice had faster basal heart rate, and ventricular tachycardia upon exposure to 2% isoflurane, which could be relieved by dantrolene. Basal sinus tachycardia and ventricular ectopic electrical triggering also occurred in Casq1 -KO hearts ex vivo . Accordingly, the ventricular cardiomyocytes from Casq1 -CKO mice displayed dantrolene-sensitive increased Ca 2+ waves and diastole premature Ca 2+ transients/oscillations upon isoflurane. NRVMs with Casq1-knockdown had enhanced spontaneous Ca2+ sparks/transients upon isoflurane, while cells over-expressing Casq1 exhibited decreased Ca2+ sparks/transients that were absent in cells with truncation of 9 amino acids at the C-terminus of Casq1. Structural evaluation showed that most of the Casq1 protein was present as a polymer and physically interacted with RyR2 in the ventricular SR. The Casq1 isoform was also expressed in human myocardium. Mechanistically, exposure to 2% isoflurane or heating at 41ºC induced Casq1 oligomerization in mouse ventricular and skeletal muscle tissues, leading to a reduced Casq1/RyR2 interaction and increased RyR2 activity in the ventricle. Conclusions: Casq1 is expressed in the heart, where it regulates SR Ca 2+ release and heart rate. Casq1 deficiency independently causes MH/EHS-like ventricular arrhythmia by trigger-induced Casq1 oligomerization and a relief of its inhibitory effect on RyR2-mediated Ca 2+ release, thus revealing a new inherited arrhythmia and a novel mechanism for MH/EHS arrhythmogenesis.

Selective toxic effects of tamoxifen on osteoclasts: comparison with the effects of oestrogen

1996 ◽  
Vol 149 (3) ◽  
pp. 503-508 ◽  
Author(s):  
T R Arnett ◽  
R Lindsay ◽  
J M Kilb ◽  
B S Moonga ◽  
M Spowage ◽  
...  
Keyword(s):  
Mononuclear Cells ◽  
Time Lapse ◽  
Bone Cell ◽  
Neonatal Rat ◽  
Pit Formation ◽  
Cell Counts ◽  
Bone Cell Cultures ◽  
Dying Cells

Abstract We investigated the actions of the trans- and cis-isomers of tamoxifen on the function of neonatal rat osteoclasts in vitro. Both compounds inhibited resorption pit formation by osteoclast-containing mixed bone cell cultures incubated for 24 h on cortical bone slices. Cell counts revealed that the inhibition was closely related to a cytotoxic effect, to which osteoclasts appeared particularly sensitive. Partial inhibition of resorption was seen in the presence of 2 μm trans-tamoxifen, whereas complete abolition of resorption and osteoclast viability occurred with 10 μm trans-tamoxifen; survival of mononuclear cells was unimpaired at either concentration. Cis-tamoxifen appeared to be slightly more toxic, with significant inhibitions of osteoclast viability and thus resorption pit formation at a concentration of 2 μm, and also of mononuclear cell numbers at 10 μm. Time-lapse video observations indicated that osteoclast death occurred rapidly (within 2–3 h) following exposure to 10 μm of either trans-tamoxifen or cis-tamoxifen. The morphological appearance of the dying cells was consistent with apoptosis. These results may help to explain the anti-resorptive action of tamoxifen seen in vivo in rats and humans. In contrast, oestradiol-17β consistently exerted no significant effects on resorption pit formation by rat osteoclasts over 24 h, even at grossly supraphysiological concentrations (up to 10 μm). Journal of Endocrinology (1996) 149, 503–508

scholarly journals Actomyosin-based Retrograde Flow of Microtubules in the Lamella of Migrating Epithelial Cells Influences Microtubule Dynamic Instability and Turnover and Is Associated with Microtubule Breakage and Treadmilling

1997 ◽  
Vol 139 (2) ◽  
pp. 417-434 ◽  
Author(s):  
Clare M. Waterman-Storer ◽  
E.D. Salmon
Keyword(s):  
Epithelial Cells ◽  
Dynamic Instability ◽  
Leading Edge ◽  
Unknown Origin ◽  
Time Lapse ◽  
Lung Epithelial Cells ◽  
Retrograde Flow ◽  
Microtubule Dynamic ◽  
Lung Epithelial ◽  
Time Lapse Imaging

We have discovered several novel features exhibited by microtubules (MTs) in migrating newt lung epithelial cells by time-lapse imaging of fluorescently labeled, microinjected tubulin. These cells exhibit leading edge ruffling and retrograde flow in the lamella and lamellipodia. The plus ends of lamella MTs persist in growth perpendicular to the leading edge until they reach the base of the lamellipodium, where they oscillate between short phases of growth and shortening. Occasionally “pioneering” MTs grow into the lamellipodium, where microtubule bending and reorientation parallel to the leading edge is associated with retrograde flow. MTs parallel to the leading edge exhibit significantly different dynamics from MTs perpendicular to the cell edge. Both parallel MTs and photoactivated fluorescent marks on perpendicular MTs move rearward at the 0.4 μm/min rate of retrograde flow in the lamella. MT rearward transport persists when MT dynamic instability is inhibited by 100-nM nocodazole but is blocked by inhibition of actomyosin by cytochalasin D or 2,3-butanedione–2-monoxime. Rearward flow appears to cause MT buckling and breaking in the lamella. 80% of free minus ends produced by breakage are stable; the others shorten and pause, leading to MT treadmilling. Free minus ends of unknown origin also depolymerize into the field of view at the lamella. Analysis of MT dynamics at the centrosome shows that these minus ends do not arise by centrosomal ejection and that ∼80% of the MTs in the lamella are not centrosome bound. We propose that actomyosin-based retrograde flow of MTs causes MT breakage, forming quasi-stable noncentrosomal MTs whose turnover is regulated primarily at their minus ends.

scholarly journals KCa3.1 Transgene Induction in Murine Intestinal Epithelium Causes Duodenal Chyme Accumulation and Impairs Duodenal Contractility

2019 ◽  
Vol 20 (5) ◽  
pp. 1193 ◽  
Author(s):  
Marta Sofía Valero ◽  
Mariano Ramón-Gimenez ◽  
Javier Lozano-Gerona ◽  
Pablo Delgado-Wicke ◽  
Pilar Calmarza ◽  
...  
Keyword(s):  
Intestinal Epithelium ◽  
Chloride Secretion ◽  
Nuclear Antigen ◽  
Immune Reactivity ◽  
Water Salt ◽  
Duodenal Epithelium ◽  
And Function ◽  
Spontaneous Contractions ◽  
The Impact

Abstract: The epithelial intermediate-conductance calcium/calmodulin-regulated KCa3.1 channel is considered to be a regulator of intestine function by controlling chloride secretion and water/salt balance. Yet, little is known about the functional importance of KCa3.1 in the intestinal epithelium in vivo. Our objective was to determine the impact of epithelial-specific inducible overexpression of a KCa3.1 transgene (KCa3.1+) and of inducible suppression (KCa3.1−) on intestinal homeostasis and function in mice. KCa3.1 overexpression in the duodenal epithelium of doxycycline (DOX)-treated KCa3.1+ mice was 40-fold above the control levels. Overexpression caused an inflated duodenum and doubling of the chyme content. Histology showed conserved architecture of crypts, villi, and smooth muscle. Unaltered proliferating cell nuclear antigen (PCNA) immune reactivity and reduced amounts of terminal deoxynucleotide transferase mediated X-dUTP nick end labeling (TUNEL)-positive apoptotic cells in villi indicated lower epithelial turnover. Myography showed a reduction in the frequency of spontaneous propulsive muscle contractions with no change in amplitude. The amount of stool in the colon was increased and the frequency of colonic contractions was reduced in KCa3.1+ animals. Senicapoc treatment prevented the phenotype. Suppression of KCa3.1 in DOX-treated KCa3.1− mice caused no overt intestinal phenotype. In conclusion, inducible KCa3.1 overexpression alters intestinal functions by increasing the chyme content and reducing spontaneous contractions and epithelial apoptosis. Induction of epithelial KCa3.1 can play a mechanistic role in the process of adaptation of the intestine.

scholarly journals Plasticity of basal cells during postnatal development in the rat epididymis

Reproduction ◽  
2013 ◽  
Vol 146 (5) ◽  
pp. 455-469 ◽  
Author(s):  
Winnie W C Shum ◽  
Eric Hill ◽  
Dennis Brown ◽  
Sylvie Breton
Keyword(s):  
Epithelial Cells ◽  
Postnatal Development ◽  
Tight Junctions ◽  
Vas Deferens ◽  
Structural Plasticity ◽  
Basal Cells ◽  
Cytokeratin 5 ◽  
Rat Epididymis ◽  
Immunofluorescence Labeling

Our previous study has shown that basal cells sense luminal factors by forming a narrow body projection that can cross epithelial tight junctions. As a first step toward characterizing the structural plasticity of basal cells, in this study, we followed their appearance and morphology in the rat epididymis and vas deferens (VD) during postnatal development and examined their modulation by androgens in adulthood. Immunofluorescence labeling for cytokeratin 5 showed that basal cells are absent at birth. They progressively appear in a retrograde manner from the VD and cauda epididymis to the initial segments during the postnatal weeks PNW1–3. At the onset of differentiation, basal cells are in contact with the lumen and their nucleus is located at the same level as that of adjacent epithelial cells. Basal cells then position their nucleus to the base of the epithelium, and while some are still in contact with the lumen, others have a ‘dome-shaped’ appearance. At PNW5–6, basal cells form a loose network at the base of the epithelium, and luminal-reaching basal cells are rarely detected. The arrival of spermatozoa during PNW7–8 did not trigger the development of projections in basal cells. However, cells with a narrow luminal-reaching projection began to reappear between PNW8 and PNW12 in the corpus and the cauda. Treatment with flutamide from PNW10 to PNW12 significantly reduced the number of luminal-reaching basal cell projections. In summary, basal cells exhibit significant structural plasticity during differentiation. Fewer apical-reaching projections were detected after flutamide treatment in adulthood, indicating the role of androgens in the luminal-sensing function of basal cells.

scholarly journals Uncoated Endocytic Vesicles Require the Unconventional Myosin, Myo6, for Rapid Transport through Actin Barriers

2004 ◽  
Vol 15 (5) ◽  
pp. 2253-2263 ◽  
Author(s):  
Laura Aschenbrenner ◽  
Samia N. Naccache ◽  
Tama Hasson
Keyword(s):  
Epithelial Cells ◽  
Motor Activity ◽  
Time Lapse ◽  
Early Endosome ◽  
Physical Barrier ◽  
Unconventional Myosin ◽  
Time Lapse Microscopy ◽  
Rapid Transport ◽  
Endocytic Vesicles

After clathrin-mediated endocytosis, clathrin removal yields an uncoated vesicle population primed for fusion with the early endosome. Here we present the first characterization of uncoated vesicles and show that myo6, an unconventional myosin, functions to move these vesicles out of actin-rich regions found in epithelial cells. Time-lapse microscopy revealed that myo6-associated uncoated vesicles were motile and exhibited fusion and stretching events before endosome delivery, processes that were dependent on myo6 motor activity. In the absence of myo6 motor activity, uncoated vesicles remained trapped in the actin mesh, where they exhibited Brownian-like motion. Exit from the actin mesh occurred by a slow diffusion-based mechanism, delaying transferrin trafficking to the early endosome. Expression of a myo6 mutant that bound tightly to F-actin produced immobilized vesicles and blocked trafficking. Depolymerization of the actin cytoskeleton rescued this block and specifically accelerated transferrin delivery to the early endosome without affecting earlier steps in endocytosis. Therefore actin is a physical barrier impeding uncoated vesicle trafficking, and myo6 is recruited to move the vesicles through this barrier for fusion with the early endosome.

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